Many higher organisms have adapted to environmental stresses such as extremes of temperature, dehydration, and salt solutions concentrations of urea and their adaptations has involved intracellular accumulations of small organic solutes called 'osmolytes.' Osmolytes stabilize proteins and other cell components against the extreme conditions and do so without significantly altering the biological activity of the macromolecule. The long-term goals of our research are two-fold: (I) to understand the mechanisms by which natural occurring osmolytes protein proteins under deleterious structural effects brought about by denaturing environmental stresses, and (II) to understand the mechanisms by which the osmolytes ensure appropriate functional activity of proteins in the face of these mechanisms by which the osmolytes ensure appropriate functional activity of proteins in the face of these stresses. The hypothesis that osmolyte-induced shifts in the native state ensemble are responsible for their appropriate effects on function will be investigated. Previously, we found that osmolytes interact unfavorably with the peptide backbone of proteins and this fundamental property of osmolytes is responsible for the stabilization of proteins against the denaturing environmental stresses. We will study in more detail the basic properties of the peptide backbone in solution, as well as osmolytes of kidney, and evaluate their contributions to the Free Energy of transfer of structured peptides/proteins from water to osmolyte solutions, Two experimental metrics were identified that quantifies the ability of osmolytes to force proteins to folder and contract or expand the denatured ensemble. These metrics will be developed and used to test factors believed to be important in osmolyte action of proteins, including macromolecular crowding. The marked effects protecting osmolytes have on protein folding kinetics will be explored and contrasted with effects commonly observed in the presence of the non-protecting osmolyte urea. Medically, the osmolytes have protective roles in human kidney and are of value in pharmaceutical formulations of protein drugs.